Cyclin C stimulates ß-cell proliferation in rat and human pancreatic ß-cells.

Activation of pancreatic ß-cell proliferation has been proposed as an approach to replace reduced functional ß-cell mass in diabetes. Quiescent fibroblasts exit from G0 (quiescence) to G1 through pRb phosphorylation mediated by cyclin C/cdk3 complexes. Overexpression of cyclin D1, D2, D3, or cyclin E induces pancreatic ß-cell proliferation. We hypothesized that cyclin C overexpression would induce ß-cell proliferation through G0 exit, thus being a potential therapeutic target to recover functional ß-cell mass. We used isolated rat and human islets transduced with adenovirus expressing cyclin C. We measured multiple markers of proliferation: [(3)H]thymidine incorporation, BrdU incorporation and staining, and Ki67 staining. Furthermore, we detected ß-cell death by TUNEL, ß-cell differentiation by RT-PCR, and ß-cell function by glucose-stimulated insulin secretion. Interestingly, we have found that cyclin C increases rat and human ß-cell proliferation. This augmented proliferation did not induce ß-cell death, dedifferentiation, or dysfunction in rat or human islets. Our results indicate that cyclin C is a potential target for inducing ß-cell regeneration.

Increased Aß production prompts the onset of glucose intolerance and insulin resistance.

Type 2 diabetes (T2D) mellitus and Alzheimer's disease (AD) are two prevalent diseases with comparable pathophysiological features and genetic predisposition. Patients with AD are more susceptible to develop T2D. However, the molecular mechanism linking AD and T2D remains elusive. In this study, we have generated a new mouse model to test the hypothesis that AD would prompt the onset of T2D in mice. To test our hypothesis, we crossed Alzheimer APPswe/PS1dE9 (APP/PS1) transgenic mice with mice partially deficient in leptin signaling (db/+). Body weight, plasma glucose, and insulin levels were monitored. Phenotypic characterization of glucose metabolism was performed using glucose and insulin tolerance tests. ß-Cell mass, islet volume, and islet number were analyzed by histomorphometry. APP/PS1 coexpression in mice with intact leptin receptor signaling did not show any metabolic perturbations in glucose metabolism or insulin sensitivity. In contrast, APP/PS1 coexpression in db/+ mice resulted in nonfasting hyperglycemia, hyperinsulinemia, and hypercholesterolemia without changes in body weight. Conversely, fasting blood glucose and cholesterol levels remained unchanged. Coinciding with altered glucose metabolism, APP/PS1 coexpression in db/+ mice resulted in glucose intolerance, insulin resistance, and impaired insulin signaling. In addition, histomorphometric analysis of pancreata revealed augmented ß-cell mass. Taken together, these findings provide experimental evidence to support the notion that aberrant Aß production might be a mechanistic link underlying the pathology of insulin resistance and T2D in AD.

In vivo phosphoproteomics reveals kinase activity profiles that predict treatment outcome in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) lacks prognostic and predictive markers. Here, we use high-throughput phosphoproteomics to build a functional TNBC taxonomy. A cluster of 159 phosphosites is upregulated in relapsed cases of a training set (n = 34 patients), with 11 hyperactive kinases accounting for this phosphoprofile. A mass-spectrometry-to-immunohistochemistry translation step, assessing 2 independent validation sets, reveals 6 kinases with preserved independent prognostic value. The kinases split the validation set into two patterns: one without hyperactive kinases being associated with a >90% relapse-free rate, and the other one showing ≥1 hyperactive kinase and being associated with an up to 9.5-fold higher relapse risk. Each kinase pattern encompasses different mutational patterns, simplifying mutation-based taxonomy. Drug regimens designed based on these 6 kinases show promising antitumour activity in TNBC cell lines and patient-derived xenografts. In summary, the present study elucidates phosphosites and kinases implicated in TNBC and suggests a target-based clinical classification system for TNBC.

Ghrelin's Effects on Proinflammatory Cytokine Mediated Apoptosis and Their Impact on ß-Cell Functionality.

Ghrelin is a peptidic hormone, which stimulates cell proliferation and inhibits apoptosis in several tissues, including pancreas. In preclinical stage of type 1 diabetes, proinflammatory cytokines generate a destructive environment for ß-cells known as insulitis, which results in loss of ß-cell mass and impaired insulin secretion, leading to diabetes. Our aim was to demonstrate that ghrelin could preserve ß-cell viability, turnover rate, and insulin secretion acting as a counter balance of cytokines. In the present work we reproduced proinflammatory milieu found in insulitis stage by treating murine cell line INS-1E and rat islets with a cytokine cocktail including IL-1ß, IFNγ, and TNFα and/or ghrelin. Several proteins involved in survival pathways (ERK 1/2 and Akt/PKB) and apoptosis (caspases and Bcl-2 protein family and endoplasmic reticulum stress markers) as well as insulin secretion were analyzed. Our results show that ghrelin alone has no remarkable effects on ß-cells in basal conditions, but interestingly it activates cell survival pathways, downregulates apoptotic mediators and endoplasmic reticulum stress, and restores insulin secretion in response to glucose when beta-cells are cytokine-exposed. These data suggest a potential role of ghrelin in preventing or slowing down the transition from a preclinical to clinically established diabetes by ameliorating the effects of insulitis on ß-cells.

Epoxypukalide induces proliferation and protects against cytokine-mediated apoptosis in primary cultures of pancreatic ß-cells.

There is an urgency to find new treatments for the devastating epidemic of diabetes. Pancreatic ß-cells viability and function are impaired in the two most common forms of diabetes, type 1 and type 2. Regeneration of pancreatic ß-cells has been proposed as a potential therapy for diabetes. In a preliminary study, we screened a collection of marine products for ß-cell proliferation. One unique compound (epoxypukalide) showed capability to induce ß-cell replication in the cell line INS1 832/13 and in primary rat cell cultures. Epoxypukalide was used to study ß-cell proliferation by [(3)H]thymidine incorporation and BrdU incorporation followed by BrdU/insulin staining in primary cultures of rat islets. AKT and ERK1/2 signalling pathways were analyzed. Cell cycle activators, cyclin D2 and cyclin E, were detected by western-blot. Apoptosis was studied by TUNEL and cleaved caspase 3. ß-cell function was measured by glucose-stimulated insulin secretion. Epoxypukalide induced 2.5-fold increase in ß-cell proliferation; this effect was mediated by activation of ERK1/2 signalling pathway and upregulation of the cell cycle activators, cyclin D2 and cyclin E. Interestingly, epoxypukalide showed protection from basal (40% lower versus control) and cytokine-induced apoptosis (80% lower versus control). Finally, epoxypukalide did not impair ß-cell function when measured by glucose-stimulated insulin secretion. In conclusion, epoxypukalide induces ß-cell proliferation and protects against basal and cytokine-mediated ß-cell death in primary cultures of rat islets. These findings may be translated into new treatments for diabetes.